Colloidal up-converting nanoparticles (UCNPs) are capable of converting near-infrared excitation into visible emission via mutiphoton up-conversion processes. These processes involve long-lived excited electronic states of dopant lanthanide ions—antennae (e.g. Yb3+), admixed to the crystal lattices of host materials (e.g. NaYF4). The host crystals also contain emissive lanthanide ions—emitters (e.g. Er3±). The antenna ions transfer excitation energy onto the emitter ions by way of multi-exciton annihilation/sensitization process. The overall up-conversion scheme consist of the following steps: 1) excitation of two or more antenna ions by low energy photons, 2) diffusion of the excitons mediated by energy transfer/exchange between the antenna ions, 3) formation of multi-exciton antenna/emitter complexes; 4) exciton annihilation and sensitization of excited states of the emitter ions within these complexes, followed by 4) emission of a higher energy photon. These processes, occurring at the core of the UCNP, are responsible for its photophysical properties, which are relatively insensitive to the environment. On the contrary, the periphery of the particle, which remains in constant contact with the environment, determines the UNCP biocompatibility and bio-analytical functionality.
UCNPs offer a number of advantages for biological imaging, including high photostability, narrow emission bands, near infrared excitation wavelengths for depth-resolved imaging and potentially reduced risk of photodamage. Compared to conventional multiphoton excitation, sequential absorption of photons by UCNPs occurs via population of real as opposed to virtual states; and, therefore, multiphoton excitation cross-sections of UCNPs are much higher than those of regular organic dye molecules. In fact, multiphoton excitation and emission of UCNPs can be induced by low power continuous wave (cw) sources. Such sources excite virtually zero auto-fluorescence and cause no photodamage.
There have been many efforts to utilize UCNPs in biological sensing. UCNPs can be effectively detected at cm's depths in the tissue, including applications in optical tomography, as well as provide opportunities for single-particle detection. Multicolor nature of UCNP emission has been recently demonstrated as a valuable tool for in vivo imaging. Nonetheless, all so far reported experiments capitalize on the advantage of near-infrared excitation to induce emission of UCNPs, whereas very few specific biological analytes have been addressed, e.g. lectin. Furthemore, in spite of the fact that the multiphoton-nature of UCNP excitation is a natural prerequizite for multiphoton microscopy applications, no high-resolution depth-resolved imaging using UCNPs has yet been performed.
UCNPs can be synthesized by a variety of methods. In the majority of these synthetic schemes, and especially in those schemes that yield the maximally emissive nanoparticles (vide infra), UCNPs are stabilized by coats of the so-called supporting ligands, such as oleic acid, polyethyleneamine etc, or even by secondary inorganic shells (e.g. mesoporous silica).
The role of this layer is to control the formation of nanocrystals, keep the UCNPs in solution during the synthesis and maximize the yield of a particular crystalline phase (β- or hexagonal phase), which is characterized by the highest up-conversion efficiency. The supporting organic ligands are attached to the UCNP surfaces by their polar head groups (carboxyls or amines). The long hydrophobic tails support crystals in non-polar solutions, but make them completely insoluble in aqueous environments. For biological applications these ligands shoul be replaced by hydrophilic coats to ensure high aqueous solubility.
Solubilization of UCNPs in aqueous environments has been attempted by several methods, including solvothermal, hydrothermal, ionothermal methods and the method of thermal decomposition. These methods differ in their efficiency and generality, but presently there is no universal approach to the synthesis of truly water-soluble UCNPs with surface groups suitable for functionalization and linking to biological targets.
Accordingly, there exits a need for improved nanoparticles that would provide enhanced solubility in water.